Need advice in primer designing

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    • #6760
      aaroncjj_2020
      Participant

      dear all

      i’ve an unpublish sequence of the coat protein of banana mild mosaic virus. i hv tried some primers but it doesnt give me the sequence that i wan. hope to get some advice in dsigning primers for my sequence.

      thanks a lot.

    • #66390
      oppox
      Participant

      its hard to know where to start without seeing the gene. the forward primer should be located in the beginning and the reverse in te end of the gene. thats pretty much it.
      Its hard to say anything but that right know since I dont know what u are going to do with it, are u having primers with a restriction site?
      what sytem are u going to put it in?
      also u can check up on touchdown PCR if u get unspecific binding.
      If your primers contain to much AT u can add some GC.

    • #67941
      lupine_nickt
      Participant

      Keep them a similar m.w. & length; at least 15 b.p., probably more (but not too many – you’re not after a megaprimer, after all); try to end the primers on a G/C.

      That’s about it, really.

      xF,

      …Nick

    • #67951
      SororSaudade
      Participant

      have you tried different annealing temperatures?

      hopefully this paper will help you as it did to me:

      http://bioinformatics.oxfordjournals.or … t/bti633v1

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